SNA Research Conference Vol Plant Pathology. Mark Windham Section Editor and Moderator. Plant Pathology Section

Transcription

1 Plant Pathology Mark Windham Section Editor and Moderator 419

2 Interaction between culitvars of Periwinkle and plant pathogenic pseudomonas C. Korsi Dumenyo and Paul Agyemang Department of Agricultural Sciences, Tennessee State University Campus Box 9610, Nashville, TN Keywords: Bacterial wilt, pseudomonas, periwinkle, vinca Significance to the Industry: Plant disease is a major contributor to losses in agriculture. Many plant disease-causing organisms enter the host tissues through the leaf surface. Plant pathogenic Pseudomonas species are ubiquitous in crop fields and cause disease on a wide range of cultivated plant species. The findings of this study can be used by growers to identify cultivars that are most susceptible to infection by these leaf surface pathogens. Nature of Work: Plant pathogenic pseudomonads make up a diverse group of bacterial plant pathogens with respect to their genetics, ecology, and pathology. Many pathovars of Pseudomonas syringae, which are normally leaf surface inhabiting epiphytes switch into pathogenic phase to cause disease [1, 2]. There are over 50 pathovars of P. syringae attacking different hosts in all climatic zones and this pathogen is arguably the most important plant pathogenic bacterium [3]. The bacterium inhabits the leaf surface together with yet uncharacterized diversity of microbes. In order to fully understand how these bacteria can interact with ornamental plant species, we have studied the effect of four strains of Pseudomonas syringe on four cultivars of periwinkle. Seedlings of periwinkle cultivars, First Kiss Blueberry, Titan Blush, Pacific Red and Mediterranean Apricot Broadeye were transplanted into Fafad Mix #2 in 4 x 4 square pots. These plants were each inoculated with P. syringae pathovars syringae 728a, P. syringae pv. tomato DC3000, P.s. pv. tabacci strain 5 and P. s. glycenia race 1. Bacteria were grown on King s Medium B agar overnight and the cells were suspended into Phosphate buffer (ph 7) to A 600 of 0.1. A randomized complete block design was used in which the bacterial strains were treated as blocks. This allowed the comparison of the cultivars within each block. There were five replicates (plants) per treatment within a block. About 50 ml of the bacterial suspension was spray-inoculated onto the five plants within the block. The inoculated plants were kept in the greenhouse and watered as needed. The plants were scored for disease severity six (6) weeks after inoculation. The severity was based on an index ranging from 0 (the plants look healthy without any apparent disease symptoms) to 5 with all leaves showing symptoms. To avoid bias, the plants were independently scored by two workers using the same severity index. Analysis was performed using the mean of the two scores for each plant. 420

3 Results and Discussion: Of the four cultivars, Pacifica Red showed the least symptoms. None of the four strains produced average disease with severity of more than 0.5. P. s. tabaci-inoculated plants showed symptoms that were even less severe than buffer-inoculated plants. The cultivar Mediterranean Apricot Broadeye and Titan Blush also had very little or no symptoms. The most severe symptom on former was incited by P.s.s 728a with an average score of 0.5 while on the latter it was Pst DC3000 with an average score of The fact that the average severity scores on many pathogen-inoculated plants were comparable to those of controlled buffer-inoculated plants suggests that these cultivars are fairly tolerant to Pseudomonas infections. The most severe symptoms were observed on First Kiss Blueberry and were incited by P. syringae pv. syringae 728a with a score of 3.05 and P. syringae pv. tomato DC3000 with a score of 0.8. These observations suggest that First Kiss Blueberry is the most susceptible to bacterial infection among the cultivars tested and could also be susceptible to infection from nonbacterial pathogens as well. Literature Cited 1. Lindow, S.E. and J.H. Leveau, Phyllosphere microbiology. Curr Opin Biotechnol, (3): p Dulla, G., et al., A Closer Look at Pseudomonas syringaeas a Leaf Colonist. ASM News, (10): p Schroth, M.N., D.C. Hilderbrand, and N.J. Panopoulos, Phytopathogenic Pseudomonads and Related Plant-Associated Pseudomonads, in The Prokaryotes. 2000, Springer-Verlag. 421

4 Figure 1. Symptoms on the leaves of Periwinkle cv. First Kiss Blueberry inoculated with P. syringae pv. syringae 728a (right 3 leaves) and buffer (left 3 leaves). The plants were spray inoculated with bacterial suspension and kept in greenhouse under natural light. 422

5 Figure 2. Disease severity of four Periwinkle (Vinca, Catharanthus) inoculated with four strains of Pseudomonas syringae pathovars. 423

6 Figure 3. A healthy (left) and diseased (right) plant of Periwinkle cultivar First Kiss Blueberry. The plant on the left was inoculated with buffer and the one on the right was inoculated with P. syringae pv. syringae 728a. 424

7 Anthracnose: a new disease in ornamental grasses Yonghao Li 1, Mark Windham 1, Robert Trigiano 1, Phillip Wadl 1, Kevin Moulton 1, Alan Windham 2 and James Spiers 3 1 Dept. of Entomology and Plant Pathology, University of Tennessee Knoxville, TN Soil, Plant and Pest Center, University of Tennessee, Nashville, TN Thad Cochran Southern Horticultural Laboratory, Poplarville, MS mwindham@utk.edu Index Words: switchgrass, Panicum virgatum, anthracnose, Colletotrichum graminicola Significance to Industry: Switchgrass (Panicum virgatum L.) is one of most graceful native perennial grasses; it is a very popular ornamental plant in landscapes because it requires minimum inputs of fertilizer and considered to be resistant to disease and insects. Because of expansion of interest and demand for ornamental grasses in landscapes, millions and millions of units of grasses are produced annually in nurseries and distributed throughout the United States. Outbreak of infectious diseases in densely produced grasses in nurseries and introduction of new diseases through the distribution of diseased grasses are potential threats to flourishing nursery and landscape industries. Survey and identification of diseases are important steps to prevent disease outbreaks and protect switchgrass production growing. In this paper, we reported a new disease - anthracnose of switchgrass and describe disease symptoms and the causal agent. Nature of Work: Switchgrass is a native perennial warm season grass that is used primarily for soil conservation, forage production, ornamental plantings and more recently as a biomass crop for ethanol production. Severe leaf spot symptoms were observed on ornamental switchgrass Prairie Sky plants in the summer of Symptoms were observed as oval to fusiform brown lesions with acervuli and black setae in necrotic centers. The objective of this study was to identify the pathogen of the leaf spot disease in switchgrass Sections of diseased areas were stained with 0.05% Cotton Blue for the observation of acervuli, setae and conidia. Fungal isolation was conducted by disinfesting small sections of typical lesions using 10% Clorox (5.3% NaCIO) for 2 min, rinsing in sterilized water three times and placing on half-strength potato dextrose agar (PDA). The isolated fungal culture was suspended in sterilized water and spread on 1% water agar plates. Colonies arising from single spores were transferred to half-strength PDA plates to obtain single spore isolates. Fungal cultures were incubated at 77F (25C) and colony characteristics were described 5 days after incubation. Spore germination and appressoria formation were observed by dropping conidia suspension on plastic sideslips after 24 h incubation. Pathogenicity of isolates was confirmed by spraying a conidia 425

8 suspension (10 5 spores/ml) on the adaxial surface of detached leaf segments using a mini sprayer. Incubation took place in a transparent plastic box with two layers of moist filter paper at room temperature with 12 h photoperiod. Results and Discussion: Conidia were hyaline, falcate, fusiform and range from µm long 3-6µm wide. Setae were black, acicular with 3-5 septate, ranging from µm long. Conidia germinated and formed appressoria at the end of short germ tubes in drops of water on plastic cover slips 20 h after incubation. Appressoria were oval or pear shaped and ranged from 8-12 µm long and 5-7 µm wide. Fungal colonies on halfstrength PDA were diffuse with irregular margin. Aerial mycelium was fluffy and light-gray color. Conidiomata with black setae formed in rings on white vegetative mycelia at 2 days after inoculation. These morphological characteristics are consistent with the description of Colletotrichum graminicola (Cesati) G.W. Wilson (1). Symptoms, observed 5 days after inoculation, were characterized by necrotic spots, and then expended as fusiform redbrown lesions with acervuli formation in the gray lesion center. The fungus isolated from infected leaves showed same morphological characteristics of the isolates previously inoculated. To our knowledge, this is the first report of C. graminicola infecting switchgrasses in the United States. Literature Cited: 1. Sutton, B. C The Coelomycetes: Fungi imperfecti with pycnidia, acervuli and stromata. Commonwealth Mycological Institute, Kew, Surrey, England. 426

9 Resistance to rust in hydrangeas Yonghao Li 1, Mark Windham 1, Robert Trigiano 1, Alan Windham 2, Sandra Reed 3, James Spiers 4, and Timothy Rinehart 4 1 Dept. of Entomology and Plant Pathology, University of Tennessee Knoxville, TN Soil, Plant and Pest Center, University of Tennessee, Nashville, TN USDA/ARS Floral & Nursery Plants Research Unit, McMinnville, TN Thad Cochran Southern Horticultural Laboratory, Poplarville, MS mwindham@utk.edu Index Words: Pucciniastrum hydrangeae, Hydrangea arborescens, rust, cultivar, resistance Significance to Industry: Smooth hydrangea, Hydrangea arborescens L., is a group of native shrubs in the eastern North America. Compared to the hardness zones 6-9 for bigleaf hydrangea H. macrophylla, H. arborescens has wide range adaptability from zone 3 to zone 9. The mature and young leaves in spring are also more frost-resistant than those of H. macrophylla and H, serrata (1). In 2008 and 2009, severe rust disease was observed on H. arborescens Annabelle in the trial garden on the University of Tennessee campus (Fig. 1). In the present paper, we report a highly resistant cultivar, Frosty, to rust disease according to comparison of uredium numbers and sporulation in vitro among seven H. arborescens cultivars. These results provide important information for breeders to develop smooth hydrangea cultivars with rust resistance and for landscape designers and nurserymen to select smooth hydrangea for gardens and nursery production. Nature of Work: Genetic resistance is an important tool in the integrated pest management of plant diseases. In order to understand resistance to rust in smooth hydrangea, seven cultivars, Annabelle, Frosty, Green Dragon, Mayes Starburst, Pink Pincushion, Ryan Gainey, and White Dome of H. arborescens were evaluated for colony numbers and sporulation in vitro. Leaf disks (1 cm diameter) were placed abaxial surface up onto moist paper towels in plastic boxes. After inoculation in a settling tower, boxes were sealed with parafilm and incubated at 72 F( 22ºC) with 12 h photoperiod. Ten days after inocualtion, leaf disks were fixed and cleared with 0.15% trichloroacitic acid in chloroform-alcohol solution for 2 days and softened in distilled water over night. Cleared leaf disks were embedded in 50% general glycerol, covered with glass slips and sealed with permount. Slides were examined under a stereo microscope for assessing uredium numbers on each leaf disk. For urediniospore production, leaf disks were sampled at 10 days after inoculation and vortexed in 1.5 ml eppendorf tubes with 1 ml 0.1% Tween 20 solution for 30 sec. Tubes were centrifuged 5 min at 10,000 rpm/min, supernatant was removed, and pellets were resuspended in 100 µl distilled water. Urediniospores were 427

10 assessed by counting spore numbers using a hemocytometer under a compound microscope. The experiments were arranged in a randomized complete block design with three subsamplings (disks) in each treatment (cultivar) and block. Analysis of variation and comparison of means at 0.05% of the least significant difference were conducted using SAS software. Results and Discussion: All seven H. arborescens cultivars tested were infected by P. hydrageae and formed uredia and urediniospores. However, significant differences in uredium numbers and urediniospore production per leaf disk were detected among cultivars (Fig. 2). The greatest and least numbers of uredia/disk were found on Green Dragon and Frosty, respectively. Intermediate reactions were detected on the other cultivars, in which Mayes Starbust and Annabelle had significantly greater numbers of uredia than Ryan Gainey, Pink Pincusion and White Dome. The results of comparison of spore production showed that cultivar Frosty had the lowest numbers of urediniospores/cm 2 among the seven cultivars. Differences in spore production among the other cultivars were not detected with the exception of significant higher value of sporulation for Ryan Gainey than that for Pink Pincushion. The results of the present study revealed the variation in resistance to rust in P. arborescens and a higher level of resistance in cultivar Frosty with lower uredium formation and urediniospore production. In the other six P. arborescens cultivars, differences in numbers of uredia/disk were not associated with the differences in sporulation. The reason for this might be the difference in sporulation for uredia that were in various size and mature stages. An investigation of differences in latent period and changes in time of sporulation among cultivars could provide more information on resistance components other than uredium and uredidiospore numbers on a unit leaf area. Literature Cited: 1 Dirr, M Hydrangeas for American Gardens. Timber Press, Inc., Portland, OR. 428

11 Figure 1. Diseased Hydrangea arborescens Annabelle showing rust symptoms in a landscape 400 a 3.0E+03 Uredia number/disk Spores/cm2 Uredium numbers/disk ab b ab b ab c a c b d ab 2.0E E+03 Urediniospores/disk 0 Green Dragon Hayes Starbust Annabelle Ryan Gainey Pink Pincushion White Dome e c Frosty 0.0E+00 Figure 2. Comparisons of uredium numbers and urediospore numbers per leaf disk, respectively, among seven smooth hydrangea cultivars. Bars with the same letter represent the difference was not significant. 429

12 Search for the No Spray Rose Continued A. K. Hagan 1 and J. R. Arkidge 2 1 Department of Entomology and Plant Pathology, Auburn University, Auburn, AL Brewton Agricultural Research Unit, Brewton, AL Key Words: black spot, Cercospora leaf spot, disease resistance, Diplocarpon rosae, Cercospora rosicola Significance to Nursery Industry: In a simulated landscape setting, newly released shrub rose varieties differed considerably in their susceptibility to black spot and Cercospora leaf spot. Whereas, Blushing Knock Out, Double Knock Out, Knock Out, and Pink Knock Out were nearly immune to black spot and Cercospora leaf spot, all suffered some non-disease related leaf yellowing and premature defoliation. In contrast to Rainbow Knock Out and to a lesser extent Coral Drift, protective fungicides probably would not be required to control Cercospora leaf spot in nursery and landscape plantings of Peach Drift, Ivory Drift, Red Drift, Lovely Fairy, Home Run, and Palmengarten Frankfort. Roses with partial resistance to black spot included Pink Drift, Belinda s Dream, Bonanza, and White Meidiland. Eureka, Gourmet Popcorn, Rabble Rouser, Rockin Robin, Julia Child, About Face, Hot Cocoa, Heart N Soul, Baby Love, and Johann Strauss proved highly susceptible to black spot and would require an intensive fungicide program to maintain their health and beauty in the landscape. Nature of Work: In Alabama s often wet and warm climate, diseases pose a significant threat to the health and beauty of roses. Black spot is the most common and destructive disease on roses. On black spot-susceptible roses, leaf spotting and early defoliation, which may appear as early as mid-april, will intensify until the plants are totally defoliated. In addition to being unattractive, defoliated roses also produce fewer blooms and greatly reduced shoot growth. To maintain the health and beauty of susceptible roses, weekly fungicide applications starting at leaf-out and continuing through the first hard frost are required. Black spot is not the only disease that commonly damages Alabama s roses. Cercospora leaf spot also causes a noticeable leaf spotting and premature defoliation on some roses, particularly on increasingly popular shrub roses. Typically, defoliation levels on Cercospora leaf spot-damaged roses are not as high as those damaged by black spot. Left unchecked, Cercospora leaf spot may slow rose growth nearly as much as black spot. Fungicides applied to control black spot will also protect roses from Cercospora leaf spot. While the powdery mildew is considered nationally second in importance to black spot, damaging outbreaks of this disease are sporadic in Alabama. When this disease does appear, signs and symptoms of powdery mildew may be seen in early to mid-spring and 430

13 again in late fall. Due to the low risk damage from this disease, fungicides targeting powdery mildew are rarely needed. Shrub roses are loosely defined group of unrelated heirloom, garden, Floribunda, and other modern hybrid roses that are marketed as having superior hardiness and vigor compared with the classic hybrid tea rose. Their growth habits range from erect bushy to sprawling, low-growing ground covers forms with multiple simple, semi-double, or double blooms on each stem. Popularity of shrub roses is due in part to claims of resistance to often unspecified diseases, and the possibility of eliminating or greatly reducing the number of protective fungicide treatments needed to control diseases that plague roses. The objective of this study was to determine the susceptibility of newly released shrub rose selections to black spot, Cercospora leaf spot, and powdery mildew in a simulated landscape planting. On 14 February 2008, bare-root roses were transplanted into raised beds on 8 foot centers with 13 feet between rows at the Brewton Agricultural Research Unit in Brewton, AL. A drip irrigation system was installed shortly after planting and the beds were mulched with aged pine bark. After each rose was pruned, fresh aged pine bark was distributed on 18 February An application of 2.1 oz of analysis fertilizer per plant on 14 April was followed by 16 July and 7 August applications of 1.8 oz per plant of a analysis fertilizer. A split plot design consisting of five replications with rose varieties as the main plot and fungicide treatment as the sub-plot was used. Daconil Weather Stik 6F fungicide at 2 quarts per acre was applied at 2- and 4-week intervals to one plant in each plot. The remaining plant in each plot was not treated with the above fungicide. Fungicides were applied with a hand wand with a TX-8 nozzle to run-off using a tractor-mounted sprayer from 16 April to 7 October. Black spot (BS) and Cercospora leaf spot (CLS) damage was visually rated on 4 September using a modified 1 to 10 Florida peanut leaf spot rating scale where 1 = no disease, 2 = very few lesions in canopy, 3 = few lesions noticed in lower and upper canopy, 4 = some leaf spotting and < 10% defoliation, 5 = lesions noticeable and < 25% defoliation, 6 = lesions numerous and < 50% defoliation, 7 = lesions very numerous and < 75% defoliation, 8 = numerous lesions on few remaining leaves and <90% defoliation, 9 = very few remaining leaves covered with lesions and < 95% defoliation, and 10 = plants defoliated. Significance of treatment effects were tested by analysis of variance and the least significant difference (LSD) test (P<0.05). Results and Discussion: In 2009, rainfall totals for the months of March, April, May, July, August, September and October were average to above average, while totals for June fell below the 30-yr average for the study location. As a result of frequent showers, black spot and Cercospora leaf spot ratings for many rose varieties were higher in 2009 compared with the previous year. Results for the non-fungicide-treated roses are presented in Table 1. Overall, the level of leaf spotting and premature defoliation was heavier for black spot compared with 431

14 Cercospora leaf spot. No mixed outbreaks of black spot and Cercospora leaf spot occurred on any rose variety. Of the varieties screened, only Knock Out, Blushing Knock Out, and Pink Knock Out were free of both black spot and Cercospora leaf spot (Table 1). Among the remaining cultivars not damaged by Cercospora leaf spot, Pink Drift and Belinda s Dream showed the highest level of black spot resistance. Moderate levels of black spot-related leaf spotting with some premature defoliation were noted on the White Meidiland, and Bonanza. Rose varieties that suffered greater than 50% black spotinduced premature defoliation and heavy spotting of the remaining leaves included Eureka, Gourmet Popcorn, Rabble Rouser, Rockin Robin, Julia Child, About Face, Hot Cocoa, Heart N Soul, Baby Love, and Johann Strauss. Among all rose varieties damaged by Cercospora leaf spot, the highest level of leaf spotting and premature defoliation was noted on Rainbow Knock Out and Coral Drift. The remaining Drift series varieties except for Pink Drift, as well as Lovely Fairy, Home Run, and Palmengarten Frankfort showed no black spot symptoms but developed moderate Cercospora leaf spotrelated leaf spotting with some defoliation. Cercospora leaf spot was not seen on the black spot-damaged roses. As previously noted (2,3), shrub roses differed considerably in their reaction to black spot and Cercospora leaf spot. In addition rose varieties were damaged by either black spot or Cercospora leaf spot but not simultaneously by both diseases (2,3). On susceptible varieties, defoliation levels were usually higher with black spot compared with Cercospora leaf spot (2). In the absence of protective fungicide treatments, Knock Out, Pink Knock Out, Double Knock Out, and Blushing Knock Out remained essentially free of black spot and Cercospora leaf spot (Table 1). While similarly high levels of resistance to both of the above diseases were previously reported (1,3), premature defoliation, which apparently was not disease related, was observed in both this and the above studies. In contrast to the above Knock Out varieties, Rainbow Knock Out rose proved to be so susceptible to Cercospora leaf spot that this selection cannot be produced in a nursery or maintained in landscapes without fungicide protection. Among the Drift groundcover roses, an unacceptably high level of Cercospora leaf spot-induced defoliation was seen on Coral Drift. While light to moderate defoliation was noted on Peach Drift, Ivory Drift, and Red Drift as well as Palmengarten Frankfort and Home Run, these roses could be maintained in the landscape without protective fungicides. While Home Run proved to be more susceptible to Cercospora leaf spot than noted by Mynes et al. (3), the level of defoliation on Palmengarten Frankfort and Lovely Fairly reported in the Tennessee study did not greatly differ from that noted here. A summary of the reaction of shrub roses evaluated at Brewton to black spot, Cercospora leaf spot, and powdery mildew along with a description of the growth habit and flower color is presented in Table 2. In summary, Eureka, Gourmet Popcorn, Rabble Rouser, Rockin Robin, Julia Child, About Face, Hot Cocoa, Heart N Soul, Baby Love, Be-Bop, and Johann Strauss, which 432

15 suffered a minimum of 50% defoliation, were unacceptably susceptible to black spot. Previously, Mynes et al. (3) reported that Be-Bop and Julia Child but not About Face were susceptible to black spot. Rose selections that displayed partial resistance to black spot included Pink Drift, Belinda s Dream, Bonanza, and White Meidiland. In Tennessee (3), Belinda s Dream showed a higher level of disease than was noted in this study. Fungicide treatments may not be needed to maintain the health and vigor of the latter four roses in landscape plantings. Literature Cited: 1. Hagan, A. K. and J. R. Akridge Unknown leaf spotting and defoliation on Knock Out roses. SNA Res. Conf. 54: Hagan, A. K., M. E. Rivas-Davila, J. R. Akridge, and J. W. Olive Resistance of shrub and groundcover roses to black spot, Cercospora leaf spot, and impact of fungicide inputs on the severity of both diseases. J. Environ. Hort. 23: Mynes, J, M. Windham, A. Windham, Y. Li, W. Copes, and J. Spiers No Spray rose cultivars for the mid South. Proc. Southern Nursery Assoc. 52:

16 Table 1. Characteristics and reaction on non-fungicide treated roses to black spot and Cercospora leaf spot in a simulated landscape planting in Brewton, AL in Disease rating* Cercospora Rose Variety Black spot leaf spot Coral Drift 1.0 h** 5.8 a Ivory Drift 1.0 h 4.4 bc Peach Drift 1.0 h 5.0 b Pink Drift 3.8 g 1.0 e Red Drift 1.0 h 4.8 bc Johann Strauss 7.2 bcd 1.0 e Eureka 7.7 abc 1.0 e Blushing Knock Out 1.0 h 1.0 e Pink Knock Out 1.0 h 1.0 e Rainbow Knock Out 1.0 h 6.4 a Double Knock Out 1.0 h 1.2 e White Meidiland 5.0 f 1.0 e Lady Elsie May 6.4 e 1.0 e Knock Out 1.0 h 1.0 e Easy Going 7.2 bcd 1.0 e Julia Child 7.0 cde 1.0 e Palmengarten Frankfort 1.0 h 4.4 bc About Face 7.0 cde 1.0 e Belinda s Dream 4.4 fg 1.0 e Hot Cocoa 7.4 bcd 1.0 e Baby Love 7.4 bcd 1.0 e Gourmet Popcorn 7.6 abc 1.0 e Lovely Fairy 1.0 h 4.2 c Rabble Rouser 7.8 ab 1.0 e Rockin Robin 8.2 a 1.0 e Be-Bop 7.2 bcd 1.0 b Home Run 1.0 h 4.8 bc Bonanza 5.0 f 1.0 e Heart N Soul 7.0 cde 1.0 e Pretty Lady 6.8 de 1.0 e *Black spot and Cercospora leaf spot incidence was visually assessed using the modified Florida 1 to 10 peanut leaf spot rating scale on 4 Sep. **Means in each column that are followed by the same letter are not significantly different according to the least significant difference test (P<0.05). 434

17 Table 2. Reaction of shrub rose selections to diseases at Brewton Agricultural Research Unit. Disease reaction Cercospora leaf spot Powdery mildew Growth habit Flower color Rose selection Black spot Coral Drift HR MS R Low, Spreading Coral Orange Ivory Drift HR R HR Low, Spreading White Peach Drift HR R HR Low, Spreading Peach Apricot Pink Drift R HR HR Low, Spreading Deep Pink Red Drift HR R HR Low, Spreading Scarlet Red Johann Strass S HR HR Bush Pink Eureka S HR HR Upright, Apricot Yellow spreading Blushing Knock Out HR HR HR Bush Light to Shell Pink Pink Knock Out HR HR HR Bush Pink Rainbow Knock Out HR S MS Bush Coral Pink Double Knock Out HR HR HR Bush Deep Red Knock Out HR HR HR Bush Red White Meidiland MS HR HR Spreading White Lady Elsie May S HR HR Bush Coral Pink Easy Going S HR HR Upright Yellow Julia Child S HR HR Rounded Yellow Palmengarten HR MS HR Low, Spreading Pink Frankfort About Face S HR HR Bush Golden Orange Belinda s Dream MS HR HR Upright Pink Hot Cocoa S HR HR Bush Russet Baby Love S HR HR Upright Yellow Gourmet Popcorn S HR HR Bushy, Bright White Cascading Lovely Fairy HR R HR Bushy, Deep Pink Cascading Rabble Rouser S HR HR Bushy, Rounded Yellow/Gold Rockin Robin S HR HR Bushy, Rounded Red/White stripes Be-Bop S HR HR Bushy, spreading Cerise w/ eye Home Run R R HR Bushy, rounded Flame Red Bonanza MS HR HR Bush Yellow Heart n Soul S HR HR Rounded, spreading White edged Lipstick Red Pretty Lady MS HR HR Rounded Creamy White Key to Disease Abbreviations: S = susceptible (require fungicide applications every 1 to 2 weeks), MS = moderately susceptible (monthly fungicide applications suggested), R = resistant (no fungicides needed), HR = highly resistant (no fungicides needed). 435

18 Impact of application timing and number on the control of stem rot with Heritage fungicide A. K. Hagan 1, J. W. Olive 2 and J. Stephenson 2 1 Department of Entomology and Plant Pathology, Auburn University, Auburn, Mobile Ornamental Research Unit, Mobile, AL Key Words: Southern blight, Sclerotium rolfsii, chemical control, azoxystrobin Significance to Nursery Industry: In Alabama, stem rot is a common and often fatal disease on container-grown aucuba, hosta, and a few woody ornamentals. While strict sanitation practices can help reduce disease-related losses, fungicide drench treatments are often required to protect target container stock from stem rot. Heritage 50WDG, which is registered for the stem rot control on woody ornamentals, proved most effective when multiple drenches were made during the summer at the 4 oz/100 gal rate. Lower rates of Heritage 50WDG significantly reduced plant mortality compared with the nontreated control but failed even when applied at monthly intervals to consistently give the high level of stem rot control that was obtained with the ProStar 70W standard. Nature of Work: Stem rot, which is caused by the soil-borne fungus Sclerotium rolfsii, is a damaging and usually fatal disease of selected container-grown woody and herbaceous ornamentals (1,3). While destructive outbreaks occur most often in Alabama on aucuba and hosta, other container crops that are targets of this disease include butterfly bush, forsythia, flowering pear, and Prague viburnum. While sanitation practices such as cleaning recycled containers, clearing crop debris on production ranges, and discarding old potting media will help reduce the occurrence of stem rot in container stock, fungicide drenches are required to prevent sizable losses in susceptible crops. Stem rot is not a year round threat on container grown ornamentals. Typically, disease outbreaks occur when day and night temperatures exceed 85F and 70F, respectively. As a result, preventative fungicide drenches are needed only during the summer months when the pathogen is active and host plants are vulnerable. Previously, Hagan and Olive (2) noted that drenches of ProStar 70W gave excellent control of stem rot on aucuba. Currently, Heritage 50WDG (azoxystrobin), Medallion 50W (fluidioxonil), and ProStar 70W (flutolanil) are registered as drench treatments for the control of stem rot on container-grown ornamentals (2). Information concerning the timing and number of fungicide treatments needed to prevent stem rot outbreaks has not been determined. The objective of this study was to assess the efficacy of Heritage 50WDG rate and number of drench treatments on the control of stem rot on aucuba as compared with the industry standard ProStar 70W. Container-grown aucuba Gold Dust in #1 (C-650) containers were purchased from a nursery. Plants were maintained outdoors on a clam shell-covered bed under 47% shade 436

19 cloth and watered daily with overhead impact sprinklers. The experimental design was a randomized complete block consisting of five (5) single plant replications. Fungicides, which were applied as a drench at a volume 0.2 qt per container, were poured directly onto the surface of the pine bark:peat moss potting medium. Drench programs with Heritage 50WG at 1,2, 4, and 8 ounces/100 gallon consisted of a single application on 18 June, two applications at 6 week intervals on June 18 and August 4, and three monthly applications on June 18, July 14, and August 18. Drenches of ProStar 70W were made on the same monthly schedule as Heritage 50W. Specific application dates are listed in Table 1. Inoculum was prepared by growing S. rolfsii, which was previously isolated from aucuba, on sterile oat seed in a flask at room temperature for 1 week. On 19 June, approximately 1 gram of inoculum (20 to 25 seeds) was placed in two to three holes around the base of each plant. No colonized oat seeds were placed into the potting medium of the non-inoculated control plants. Plant survival was assessed at 2-week intervals. Plant survival on September 15 is displayed in the table. Significance of treatment effects was tested by analysis of variance and least significant difference (LSD) test (P<0.05). Results and Discussion: Significant increases in aucuba survival were obtained at all rates and drench numbers with Heritage 50WDG as well as the ProStar 70W standard when compared with the inoculated control (Table 1). With Heritage 50WDG, two drenches at the 2 ounce rate, two and three drenches at the 4 ounce rate, and the 8 ounce rate regardless of the number of drench treatments gave better stem rot control than one or two drenches of the 1 oz rate of Heritage 50WDG. With the exception of the latter 1 oz drench treatments, all remaining Heritage 50WDG drench treatments controlled stem rot as effectively as the ProStar 70W standard. At each rate of Heritage 50WDG, similar stem rot control was obtained with one, two, and three drench treatment programs. Drench rate and number had a significant impact on the efficacy of Heritage 50WDG for the control of stem rot on aucuba. Aucuba was protected from stem rot with multiple drenches of the 4 oz rate of Heritage 50WDG. Effectiveness of the latter Heritage 50WDG drench treatments was comparable to the ProStar 70W standard. At the 1, and to a lesser extent 2 oz rates, Heritage 50WDG failed to effectively prevent stem rot-related plant death. While the 8 oz rate of Heritage 50WDG, regardless of application number, proved highly effective in controlling stem rot on aucuba, use of this rate is prohibited. Literature Cited: 1. Hagan, A. K Southern blight on flowers, shrubs, and trees. AL. Coop. Ext. Sys. Cir. ANR Hagan, A. K. and J. W. Olive Assessment of new fungicides for the control of southern blight on aucuba. J. Environ. Hort. 17(2): McRitchie, J. J Southern blight on ajuga. Fla. Dept. of Agric. Cons. Serv. Plant Path. Cir

20 Table 1. Impact of Heritage 50W drench rate and number on the control of stem rot on aucuba. Drench Aucuba Fungicide treatment and rate/100 Dates Number survival (%) gal Heritage 50WDG 1 oz 18 Jun 1 50 b* Heritage 50WDG 1 oz 18 Jun, 14 Jul, b Aug Heritage 50WDG 1 oz 18 Jun, 4 Aug 2 75 ab Heritage 50WDG 2 oz 18 Jun 1 75 ab Heritage 50WDG 2 oz 18 Jun, 14 Jul, ab Aug Heritage 50WDG 2 oz 18 Jun, 4 Aug a Heritage 50WDG 4 oz 18 Jun 1 75 ab Heritage 50WDG 4 oz 18 Jun, 14 Jul, a Aug Heritage 50WDG 4 oz 18 Jun, 4 Aug a Heritage 50WDG 8 oz 18 Jun a Heritage 50WDG 8 oz 18 Jun, 14 Jul, a Aug Heritage 50WDG 8 oz 18 Jun, 4 Aug a ProStar 70W 6 oz 18 Jun, 14 Jul, a Aug Inoculated control -- 0 c Non-inoculated control a *Means in each column that are followed by the same letter are not significantly different according to the least significant difference test (P<0.05). 438

21 Phoma Leaf Spot in Bigleaf Hydrangea M.T. Mmbaga 1, Y. H. Li 2, S.M. Reed 3, R. Trigiano 2, and M.T. Windham 2 1 Tennessee State University, Otis Floyd Research Center, McMinnville, TN 2 University of Tennessee, Knoxville, TN 3 USDA/ARS Floral and Nursery Plants Research Unit, McMinnville, TN mmmbaga@tnstate.edu Index Words: ornamentals, foliar diseases, Hydrangea macrophylla Significance to Industry: Bigleaf hydrangea (Hydrangea macrophylla) is a popular flowering shrub grown in home gardens and landscapes either in full sun or in shade. Foliage diseases can have significant impact on the appearance, health and market value of hydrangeas. A total of 77 cultivars of big leaf hydrangea grown in full sun in McMinnville TN were evaluated. While some cultivars clearly did not perform well in full sun, some grew well. Those that grew well were severely devastated by foliar diseases and suffered premature severe defoliation. This paper reports on leaf spot symptoms caused by Phoma exigua Desm. This disease can easily be confused with leaf spots caused by Cercospora hydrangeae. Nature of Work: A total of 77 bigleaf hydrangea cultivars were arranged in a completely randomized block design with four replications per cultivar. Plants were fertilized with Osmocote Pro fertilizer (Scotts-Sierra Horticultural Products Co., Maryville, OH) and watered daily using drip irrigation with a five-minute overhead sprinkler spray in mid afternoon. Leaf spot/blight symptoms were observed from early May to mid October. Causal pathogens were isolated from small pieces of leaf tissue cut from the margin of lesions. Leaf tissues were sterilized in 10% commercial bleach (4% sodium hypochlorite) for 3 minutes, rinsed twice with sterile water, blotted dry on sterile tissue paper, and placed on Potato Dextrose Agar (PDA) media. Fungal colonies on PDA were sub-cultured to generate pure cultures. The pathogenicity of isolates was tested using disease-free detached leaves of three bigleaf hydrangea cultivars, Blue Bird, Forever Pink and Nikko Blue. Plugs of mycelia were cut from 14- day-old cultures and placed on one side of the half leaf divided by mid-vein and a plug of clean PDA with no fungus was placed on the other side of the leaf as a control. Disease development was monitored and lesions were measured after 14 days. In addition to the plug-inoculation on detached leaves, spay-inoculation was conducted on bigleaf hydrangea Lady in Red in a greenhouse (24± 3 C) using an atomizer and a spore suspension of spores per ml. Leaves were allowed to dry before plants were incubated at 100% relative humidity for 36 hours in a plastic cage equipped with a humidifier. The fungus was re-isolated from disease lesions to confirm pathogenicity. Fungal growth and spore production were evaluated in PDA, V-8 and water agar under three light treatments: continuous light, alternating light and dark periods (12 h of light and 439

22 12 h of dark) and continuous darkness and characterized morphologically using a compound microscope. The fungus was also grown on nylon membrane and genomic DNA was extracted from conidia and mycelium using the DNeasy Plant Mini Kit (Qiagen Inc, Valencia, CA) following the manufacturer's protocols. PCR amplifications were performed using a DNA thermal cycler following standard PCR procedures with minor modifications. Each 50 μl PCR reaction mixture consisted of 36 μl sterile ddh 2 O, 5 μl 10X PCR buffer, 3 μl MgCl 2 (25 mm), 1.5 μl dntp (10 mm total, 2.5 mm each), 1.5 μl primer each (20 ng/μl), 0.2 μl Taq polymerase (Promega) (5 U/μl), and 1.3 μl template DNA (20 ng/μl). PCR cycles consisted of an initial denaturation step at 94 C for 4 min followed by 42 cycles of 1 min at 93 C (denaturation), 1 min at 40 to 60 C (annealing), and 2 min at 72 C (extension). The annealing temperature was set based on the primer Tm and usually a less than the lower primer Tm (Tm 5) was used as the annealing temperature for the PCR reaction. An extension cycle of 72 C for 5 min was used to terminate the reaction and finally a 4 C soak. PCR products were visualized in 1.5% agarose gel in 1X TBE, stained with ethidium bromide. Two universal primers, ITS1, and ITS4 were used to amplify the ITS region of the pathogen by using standard PCR procedures. PCR products were purified using a QIA quick PCR Purification Kit (Qiagen Inc, Valencia, CA) ( following QIAquick Spin Handbook, July 2002's protocol and sequenced by the Davis Sequencing Inc. at Davis, CA ( The sequences were analyzed and compared with all sequences of ITS region in GenBank by use of BLAST ( Results and Discussion: Disease-free leaves inoculated with mycelial plugs developed dark brown necrotic lesions associated with some leaf chlorosis, while the other half leaf of control treatment with clean PDA plugs did not develop any symptoms (Fig 1). All three cultivars were susceptible to the disease with development of necrotic lesions. Spray-inoculated Lady in Red developed small reddish brown lesions (Fig 2) while Seafoam formed large lighter brown lesions and caused defoliation. Phoma exigua was isolated from necrotic lesions on plug-inoculated detached leaves and on sprayinoculated plants. Judging from the lesion sizes and defoliation, Seafoam was more susceptible than Lady in Red. It was also apparent that susceptibility affected the symptoms. Symptoms in Lady in Red were similar to those reported for Cercospora leaf spot (2). Fungal colonies began as olive greenish, and then turned into a dark color. The fungus formed abundant spores on PDA, but not on V-8 or water agar. Continuous light was most favorable for spore production, followed by alternating dark/light periods supporting moderate spore production. Very low spore production was observed in continuous darkness. Fungal growth was very poor on water agar and no spores were detected. Black pycnidia 275 to 255 μm in diameter developed on older cultures (more than 30-days-old). The pycnidia submerged in the media oozed out cream-colored spore 440

23 masses. Conidiospores were characteristically small, one-celled, hyaline, ovoid to elongate in shape, and measuring μm with an average size of 6.0 x 2.6 μm. (Fig. 3). The fungus morphological features were characteristic of Phoma spp (1, 3). DNA sequence analysis showed a close match to Phoma exigua by 99%. Although P. exigua has been reported on H. macrophylla in the United State (2, 6), its impact and significance in hydrangea aesthetic value is not clear because other pathogens such as Gloeosporium gloesporoides, Myrothecium roridum and Cercospora hydrangeae (6) were also isolated from similar symptoms. Phoma exigua was associated with severe defoliation in Italy (4). Early infection P. exigua may predispose infected plants to other opportunistic pathogens. Early defoliation observed in Seafoam in this study suggests that P. exigua could cause early defoliation in highly susceptible cultivars and impact plant growth. This study confirms that P. exigua is involved in hydrangea leaf spot/blight disease complex and causes defoliation that may be confused with that caused by better-known pathogens (5, 6). The presence of P. exigua in disease evaluation plots may result in overestimation of Cercospora leaf spot infection and influence disease resistance readings. Literature Cited 1. Barnett, H.L. and B.B. Hunter. Illustrated genera of imperfect fungi Fourth Edition. APS Press. 218 pgs. 2. Daughtrey, M. L. et al. Page 26 in: Compendium of Flowering Potted Plant Diseases. The American Phytopathological Society. St. Paul, MN, Fitton, M., and Holliday, P. No. 253 in: CMI Descriptions of Pathogenic Fungi and Bacteria. The Eastern Press Ltd. Great Britain, Garibaldi, A., Gilardi, G., Minerdi, D., and Gullino, M. L First report of leaf spot caused by Phoma exigua on Hydrangea macrophylla in Italy. Plant Disease 90: Hagan, A. K. and Mullen, J. M Diseases of Hydrangea. Alabama Cooperative Extension System, ANR Mmbaga, M. T. Windham, M. T., and Li, Y. H Fungi associated with naturally occurring leaf spots and leaf bights in Hydrangea macrophylla in McMinnville, Tennessee. Southern Nursery Association Res. Conf. Proceed. 54:

24 a. b. c. Figure 1. Lesions produced by Phoma exigua on detached leaves of Nikko Blue inoculated with mycelia plugs (a) and on Lady in Red plants spray inoculated with conidiospores (b-c). Figure 2 Phoma exigua pycnidia 1b (1c) Figure 3. Phoma exigua pycnidiospores 442

25 Myrothecium roridum in Garden Hydrangea M. T. Mmbaga Tennessee State University School of Agriculture and Consumer Sciences Otis Floyd Nursery Research Center, McMinnville TN Index Words: ornamentals, diseases, Hydrangea macrophylla Significance to Industry: Garden hydrangea (H. macrophylla) is a popular flowering shrub grown in home gardens and landscapes in full sun and in shade. Foliage diseases impact the appearance, health and market value of hydrangeas. While powdery mildew (Erysiphe polygoni), and leaf spots (Cercospora hydrangeae and Glomerella gloerisporoides anamorph: Colletotrichum gloerisporoide) are known to cause significant damage to hydrangea foliage (4,8), other lesser-known pathogens may have been responsible for the observed symptoms (7). This paper reports on Myrothecium roridum as a pathogen of hydrangeas. Nature of Work: A total of 77 hydrangea cultivars arranged in a completely randomized block design with a replication of four plants per cultivar were grown in full sun in 3 gallons containers. Plants were fertilized with Osmocote Pro fertilizer (Scotts- Sierra Horticultural Products Co., Maryville, OH) and watered daily using drip irrigation with a five-minute overhead sprinkler spray in mid afternoon. Leaf spot and leaf blight symptoms were observed beginning in early May and increased in severity over time; by mid summer, severely infected plants were 60-80% defoliated. The causal pathogen was isolated from small pieces of leaf tissue cut from the margin of the disease lesions that were surface sterilized in 10% commercial bleach (4% sodium hypochlorite) for 3 minutes, rinsed twice with sterile water, blotted dry on sterilized tissue paper and plated on 2% Potato Dextrose Agar (PDA). All isolates were sub-cultured and grown in pure cultures. Evaluation of isolates for pathogenicity was done using a detached leaf technique. Disease-free leaves of Nikko Blue and All Summer Beauty were detached and placed in clear plastic containers lined with a double layer of tissue paper and kept wet with deionized distilled water. Each half leaf was inoculated using a 5mm plug of mycelia cut from 14-day-old cultures; a sterile plug of clean PDA was placed on the other half leaf as a control treatment. Inoculated leaves were incubated at 68, 73.5 and 78.8 F (20 C, 23 C and 26 C) with a replication of four individual leaves at each temperature. The pathogen was re-isolated to complete Koch s postulate as proof for pathogenicity. The pathogen was studied under a compound microscope and identified using its morphological features. The fungus was further evaluated for virulence on 78 hydrangea cultivars using a detached leaf technique and mycelia plugs as described above. The fungus was grown on nylon membrane and genomic DNA was extracted from conidia and mycelium using the DNeasy Plant Mini Kit (Qiagen Inc., Valencia, CA) following the 443

26 manufacturer's protocol ( PCR amplification was performed. Each 50 μl PCR reaction mixture consisted of 36 μl sterile ddh 2 O, 5 μl 10X PCR buffer, 3 μl MgCl 2 (25 mm), 1.5 μl dntp (10 mm total, 2.5 mm each), 1.5 μl primer each (20 ng/μl), 0.2 μl Taq polymerase (Promega) (5 U/μl), and 1.3 μl template DNA (20 ng/μl). PCR cycles consisted of an initial denaturation step at 94 C for 4 min followed by 42 cycles of 1 min at 93 C (denaturation), 1 min at 40 to 60 C (annealing), and 2 min at 72 C (extension). A final extension cycle at 72 C for 5 min was used to terminate the reaction and finally a 4 C soak. All PCR products were visualized in 1.5% agarose gel in 1X TBE, stained with ethidium bromide. Two universal primers, ITS1, and ITS4 were used to amplify the ITS region in the pathogen by a using standard PCR procedures. PCR products were purified by use of QIA quick PCR Purification Kit (Qiagen Inc, Valencia, CA) ( following the instructions of the protocol (QIAquick Spin Handbook, July 2002) and sequenced (Davis Sequencing Inc. Davis, CA, Sequences were analyzed and compared with all sequences of ITS region in the GenBank by use of BLAST ( Results and Discussion: In H. macrophylla, leaf spot/blight characterized by brown necrotic lesions with concentric rings and ash colored centers were previously reported to be associated with anthracnose (Target leaf spot) caused by Colletotrichum gloerisporoides (4). In these studies, the fungus Myrothecium roridum was isolated from hydrangea leaf spot symptoms characterized by dark brown necrotic lesions with concentric rings and ash colored centers (Fig 1). Similar symptoms have been reported on other ornamentals such as salvia, gardenia, begonia, and New Guinea impatiens infected with Myrothecium roridum (6). These symptoms are typical of Myrothecium leaf spot (6). When disease-free detached leaves were inoculated with mycelial plugs of M. roridum, these symptoms were reproduced, while those inoculated with clean agar plugs remained symptomless (Fig 2). Initial symptoms of this disease on inoculated plants were observed beginning four days after inoculation. Infection developed at all three temperatures tested 68, 73.5 and 78.8 F. Some cultivars had larger lesions at 73.5 and 78.8 F while some had larger lesions at 68 F. M. roridum was re-isolated consistently from symptomatic tissue. These isolates produced white colonies forming dark to black spore bearing mycelia cushions (sporodochia) on the outer part of the colonies. Conidia were hyaline to slightly dark, one-celled, and ovoid to elongate, x μm in size with rounded ends. These morphological characteristics were consistent with the description of Myrothecium roridum Tode ex Fr. (1, 2, 3) and DNA sequence analysis confirmed the morphological observations (Fig 3). Out of 77 cultivars evaluated using detached leaf technique, 33 formed lesions of different sizes and 44 were not susceptible to the pathogen. Differences in levels of susceptibility were observed among the cultivars with 24 cultivars forming lesions that were more than double the size of the initial plugs. This indicated a higher level of susceptibility while nine cultivars formed small lesions. More than 150 plant species are susceptible to M. roridum. These include both ornamentals and other herbaceous and 444

27 woody plants. However, Myrothecium leaf spot has not previously been reported as a hydrangea pathogen in the United States. Although hydrangea leaf spots/blights with concentric rings have been described for anthracnose (Target leaf spot) (Colletotrichum gloerisporoides), the symptoms are also known to be typical of Myrothecium leaf spot described on other ornamentals such as salvia, gardenia, begonia, and New Guinea impatiens (4). Confirmation of pathogenicity of M roridum in hydrangea suggests a closer evaluation of disease symptoms resembling those described for Colletotrichum gloerisporoides to ensure the accuracy of the identification. Leaf spot/blight symptoms observed on hydrangea are similar to those described for M. roridum on other plants (3). Symptoms observed may be the result of a complex of the two pathogens or from either pathogens. Although we have isolated Colletotrichum sp. we have not been able to reproduce infection from those isolates. It is possible that the cultivar used for the test were resistant to the pathogen. Identification of cultivars for disease resistance requires accurate identification of the pathogen, if both pathogens occur, separate evaluations for each pathogen may be beneficial in breeding for disease resistance (7, 8). It is apparent that in middle Tennessee, hydrangea leaf spots/blights occur as a disease complex with more than one pathogen involved. Molecular methods for separating these pathogens on infected tissue will be helpful for the accurate identification of hydrangea pathogens associated with leaf spots and blights. Literature Cited: 1. Barnett, H.L. and B.B. Hunter. Illustrated genera of imperfect fungi Fourth Edition. APS Press. 218pgs. 2. Fitton, M. and P. Holliday. No. 253 in: CMI Descriptions of Pathogenic Fungi and Bacteria. The Eastern Press Ltd. Great Britain, Greenhouse Products News Magazine: October 30, Hagan, A.K. and J.M. Mullen Diseases of Hydrangea. Alabama Cooperative Extension System, ANR List.pdf 6. Mangandi, J. A., T. E. Seijo, and N. A. Peres First Report of Myrothecium roridum Causing Myrothecium Leaf Spot on Salvia spp. in the United States., Department of Plant Pathology, Gulf Coast Research and Education Center, Wimauma, FL Plant Dis. 91: Mmbaga, M. T., M.T. Windham, and Y. Li, R. and R.J. Sauvé Fungi associated with naturally occurring leaf spots and leaf bights in Hydrangea macrophylla in McMinnville, Tennessee. Southern Nursery Association Res. Conf. Proceed. 54: Mmbaga, M. T., Y. Li, R.J. Sauvé and M.T. Windham, Leaf spots and Leaf blight disease complex in Hydrangea macrophylla. Phytopathology 99:

28 Figure. 1. Leaf spot/blight symptoms concentric rings observed on hydrangea grown in full sun. Figure 2. Leaf spot/blight symptoms reproduced on detached leaves. Figure 3. Conidiospores of Myrothecium roridum. 446

29 Leaf Spot Diseases on Ornamental Flowering Cherry Margaret Mmbaga and Roger Sauve Tennessee State University, School of Agriculture and Consumer Sciences Otis L. Floyd Nursery Research Center, McMinnville, TN Additional index words: Blumeriella jaapii, Coccoomyces hiemalis, Cylindrosporium padi Significant to Industry: Ornamental flowering cherry trees (Prunus species) are closely related to orchard cherry trees that are grown primarily for their fruits. The ornamental flowering cherry is grown primarily for its springtime floral display with stunning white to pink flowers blooming in early spring before the leaves develop. Annual Cherry Blossom Festivals of Kwanzan Cherry in Washington, D.C. and Macon, Georgia display the magnificence of the flowering cherry tree. Leaf spot diseases that weaken flowering cherry trees reduce their market value, reduce plant vigor and impact plant sales by increasing the time needed for infected plants to reach prime size for sales. This report documents a leaf spot disease also known as shot hole disease that has attained economic significance to growers of flowering cherry. Nature of Work: A leaf spot problem in flowering cherry has caused increasing concern to commercial nurseries where growers reported that plant growth has been stunted for 2-3 consecutive years and tree sales to other commercial nurseries were recently rejected because of the problem. First disease symptoms were observed in early May and infected plants were immediately sprayed with common fungicides such as propiconazole (Banner Maxx) and chlorothalonil (Daconil) every two weeks. Although the two fungicides are listed for controlling cherry leaf spot, the disease continued to increase in severity and infected trees became severely defoliated and stunted in growth. It is probable that fungicide application was initiated too late after infection was established; it is also possible that the fungicides are not adequately effective. The objective of this study was to definitively identify the disease and provide grower guidance on disease management. Infected leaves were collected and placed in moist environment to induce spore formation for disease identification. Leaf debris of previously infected leaves was also placed in moist chamber to induce spore formation. Spores were observed under a compound microscope and identified using morphological features. Some spores were collected using a sterilized scalpel and were plated on 2% Potato Dextrose Agar (PDA) for direct isolation. In addition, small pieces of leaf tissue were cut from the margin of the lesions, surface sterilized in 10% commercial bleach (4% sodium hypochlorite), rinsed twice in sterile water, blot dried on sterile tissue paper and plated on PDA. The isolates 447

30 were sub-cultured to generate pure cultures and tested for pathogenicity using detached leaf technique. Disease-free leaves were placed in clear plastic containers lined with a double layer of tissue paper kept wet using sterile water. The disease free leaves were inoculated using 5mm mycelial plugs prepared from 14-day-old disease cultures. Each half leaf was inoculated with a test isolate and the other half leaf was a control treatment consisting of a sterile PDA plug. Another set of leaves were inoculated using a drop of spore suspension at 1x10 4 spores per ml and sterile water was used as control treatment. A replication of four individual leaves per isolate was used; the experiment was repeated once. Inoculated leaves were incubated at 20±3 C and monitored for disease development. The pathogen was re-isolated from necrotic lesions to complete Koch s postulate as proof for pathogenicity. The pathogenic fungus was observed under a compound microscope and identified using morphological features. Results and Discussion: Observations of the disease revealed small red-brown circular leaf spots that became necrotic and lesion centers fell out forming numerous shot holes. Infected leaves defoliated prematurely and by June, only a few flag leaves remained on most infected trees (Fig 1). Symptoms on leaf lamina were characteristically small lesions, circular, and red-brown in color with definite borders (Fig 2). New lesions appeared purplish in color before they turned red brown; the centers of the spots often dropped out producing numerous shot holes (Fig 2). These symptoms are typical of cherry leaf spot disease reported on fruit trees (4,5,6). Lesions sometimes merged to form a large irregular necrotic leaf spot. Some leaves with relatively few leaf spots and shot holes turned yellow and defoliated. By mid-july defoliation had reached 90-95% and plant growth completely ceased. Out of six cultivars that were in production, two cultivars ( Oshina and Kwanzan ) were severely affected. Two cultivars ( Snow Goose and Autumn Glory ) were moderately affected and two cultivars ( Okami, and Oxy Bonel ) did not have much infection. The susceptible cultivars had very low aesthetic value and split bark indicative of winter injury were widespread in the infested field. It is not clear whether the moderately affected Snow Goose and Autumn Glory and the more resistant Okami, and Oxy Bonel in nursery fields were more resistant to the disease or if they simply escaped infection. Evaluation of different cultivars for disease resistance will allow growers to make informed decisions on cultivars they chose to grow. A visit to another nursery that was a source of cuttings and seedlings revealed the presence of infection in the young seedlings. This observation indicated that infected cuttings were most likely the source of infection. The disease may have been introduced on a few infected plants and increased in severity, spreading over time. The presence of a large number of susceptible plants and favorable weather may have favored disease increase to economic levels. The fungus that cause this leaf spot disease produced spore bearing structures (acervuli) that were subepidermal on the host tissue and formed pinkish-whitish spore horns on the lesion centers on the underside of the leaves. Conidiospores were hyaline and filiform in shape, single-celled and some were septate. The fungus was morphologically similar to 448

31 Phloeosporella (Cylindrosporium) spp., an anamorph of B. jaapii (Syn. Coccomyces hiemalis). The leaf debris formed ascostroma (apothecia) immersed in host tissues and released ascospores that were narrower in the middle, hyaline, 1-2 celled and ellipsoid to elongate in shape. Observations on flowering cherry in the landscape revealed one landscape area that had infected trees with leaf spot and shot hole symptoms similar to those observed in the commercial nursery. Out of approximately 50 trees observed in the landscape, only two trees at one location showed the presence of the disease. The same pathogen, B. jaapii, was observed and isolated from the landscape plants. While there were four flowering cherry trees planted in that landscape, two plants near each other had severe infection and defoliated completely by mid August and the other two plants did not have any disease symptoms. The non-infected plants were separated from the infected plants by space and physical structure. It is not clear whether the disease-free plants were resistant to the disease or escaped infection due to lack of inoculum to initiate infection. It was clear that the disease was already in the landscape, but was not widespread. Control of the disease at nursery level is less complicated than at the landscape setting and it will reduce disease spread and impact on this important landscape tree. Although the affected nursery initiated fungicide applications soon after symptoms were observed, disease control was not successful. This failure in disease control could have been due to wrong choice of fungicides and/or frequent rains that interfered with fungicide sprays and efficacy. Disease control for cherry leaf spot on fruit trees typically requires five to seven fungicide applications starting from petal fall. Effective fungicides for cherry leaf spot include copper-based fungicides such as copper sulfate (Bordeaux mixture), captan and chlorothalonil, Demethylation inhibitors (DMI) and respiration inhibitors. Although fungicide applications on flowering cherry started soon after leaf spot symptoms were observed, infection may have already been widespread. On fruit trees, newly developing leaves develop infection during bloom or shortly afterwards. Reports from fruit trees indicate that the pathogen, B. jaapii overwinters on infested leaf debris and in the spring, apothecia fruiting structures develop on these leaves, releasing ascospores as primary inoculum. Ascospores are forcibly discharged and are carried upward by wind and splashing rain; the first sign of infection may be on suckers close to the ground (3). The incubation period from the first infection to the appearance of spots varies with temperature and wetting from rain and can occur in as little as five days (2,3,6,7). Repeated generations of conidiospores formed on infected leaves spread the disease throughout summer and fall. In order to determine the best time to initiate a spray program suited to Tennessee weather, it is important to monitor trees in early spring, while plants are still dormant and initiate fungicide spray program when infection is just beginning; spore traps can detect spore release and be used to improve the timing of fungicide applications. Cultural methods of controlling this disease include removal or destroying leaf litter or encouraging decomposition of leaf litter to reduce or eliminate the source of primary inoculum. It is also important to understand the disease in Tennessee weather, the source of infection in spring and the fungus mechanism of survival from one season to another. Infection of fruit trees often defoliates trees by 449

32 midsummer, weakens infected trees and reduces flowering. Repeated defoliation makes the tree more susceptible to winter injury and may eventually kill infected trees (4). The importance of this disease on flowering cherry is not known, but split barks on almost all infected trees was widespread and infected trees had very low aesthetic value. If left uncontrolled, the disease has the potential to significantly impact flowering cherry production. Literature Cited: 1. Eastwell, K.C.,G.A. Grove, D.A.Johnson, G.I. Mink. R.S. Byther R.P. Covey, and R. Parker Field guide to Sweet Cherry Diseases in Washington. Washington State University Extenssion. Publlication coe 238. EB1323E. 2. Eisenmith, S.P. and A.L.Jones A model for detecting infection periods of Coccomyces hiemalis on Sour Cherry. Phytopathology 71: McMannus, P. S., Proffer T.J., Berardi, Gruber B.R., R. Nugent, J.E. Ehret, G.R. Ma, Z. and Sundin G.W Integration of copper-based and reduced Risk fungicides for control of Blumeriella jaapii on Sour Cherry. Plant Dis. 91: Rabadoost, Mohammad Cherry leaf spot. Report on Plant Disease No September University or Illinois Extension Diagnostic Keys to Major Tree Fruit Diseases in the Mid-Atlantic Region July 3,

33 a. b. Figure 1. Branches of flowering cherry when disease free (a) and defoliation caused by leaf spot disease (b). Figs. 2. Leaf spot symptoms on flowering cherry causing shot holes 451

34 Comparison of Biorational Fungicides for Control of Black Spot on Rose J. Olive 1, A. Hagan 2, and J. Stephenson 1 1 Auburn University Orn. Hort. Research Center, Mobile, AL Department of Entomology and Plant Pathology, Auburn University Auburn, AL Index words: Diplocarpon rosae olivejw@auburn.edu Significance to Industry: Short re-entry intervals (REI) are often desired for container nursery and landscape pesticide applications and many biorational fungicides have minimal REI. Although disease pressure was low and black spot was not severe in this test, Daconil Ultrex (an industry standard), MilStop and Garden Safe Fungicide 3 reduced disease incidence when compared to the untreated control. Nature of the Work: Black spot on rose is a serious disease in both production and the landscape. In the humid South, weekly sprays are often required to maintain healthy plants with desirable foliage, greatly increasing the production and maintenance costs of growing roses. The identification of effective products for black spot control is useful to provide options for rotation of fungicides to control this disease. In the summer of 2009 miniature rose liners, Rosa x Fairy Red, were potted into 3 quart pots in a pine bark:peat moss medium (3:1 by vol) amended with 14 lb Osmocote Plus , 6 lb of dolomitic limestone, and 2 lb of gypsum per cubic yard of potting mix. Plants were maintained in full sun under overhead impact sprinkler irrigation. The experimental design was a randomized complete block design with six single plant replications. In this study, Daconil Ultrex (chlorothalonil), Liquid Copper Fungicide (copper ammonium complex, lb copper per gallon), and MilStop (potassium bicarbonate) were applied at 1 and 2 week intervals from 5 August to 30 September. The remaining fungicides: Bonide Citrus Fruit & Nut Orchard Spray Concentrate (sulfur & pyrethrins), Garden Safe Fungicide 3 (neem oil extract), Milstop + Cease (Bacillus subtillus), Bonide All Seasons Horticultural Spray Oil (petroleum oil), and Bonide Liquid Copper Fungicide (copper octanoate, copper soap) were applied at 7 day intervals from 5 August through 7 October. See Table 1 for application rates of each product. Plants were sprayed with a CO 2 pressurized sprayer. Black spot intensity was rated visually on 15 September using a modified 1 to 10 Florida peanut leaf spot rating scale where 1 = no disease, 2 = very few lesions in canopy, 3 = very few lesions noticed in upper and lower canopy, 4 = some leaf spotting and 10% defoliation, 5 = lesions noticeable and 25% defoliation, 6 = lesion numerous and 50% defoliation, 7 = lesions very numerous and 75% defoliation, 8 = numerous lesions on few remaining leaves and 90% defoliation, 9 = very few remaining leaves covered with lesions and 95% defoliation, and 10 = plants 452

35 defoliated. Significance of treatment effects were tested by analysis of variance and Fisher s protected least significant difference (LSD) test (P 0.05). Results and Discussion: Disease pressure was not severe during this evaluation. When compared with the non-treated control, significant reductions in black spot damage were obtained with weekly applications of Daconil Ultrex and MilStop as well as with Garden Safe Fungicide 3. Ratings for all other fungicide treatments in this trial were similar to the non-treated control. Table 1. Efficacy of fungicide sprays for control of black spot on miniature rose variety Fairy Red. Application Black Spot Fungicide and rate/100 gal Interval Rating* Daconil Ultrex l.4 lb. 1 wk 2.8 f** MilStop 2 lb 1 wk 3.5 ef Garden Safe Fungicide 0.8 gal 1 wk 3.8 de Daconil Ultrex 1.4 lb 2 wk 4.0 cde MilStop 2 lb 2 wk 4.2 cde MilStop 1.5 lb + Cease 1 gal 1 wk 4.2 cde Liquid Copper Fungicide 1.1 qt 2 wk 4.3 bcd Liquid Copper Fungicide 1.1 qt 1 wk 4.5 bcd Bonide Liquid Copper Fungicide 1.5 gal 1 wk 4.5 bcd Bonide Citrus, Fruit, & Nut Orchard Spray Concentrate 2 gal 1 wk 4.7 abc Non-treated control abc Bonide All Seasons Horticultural Spray Oil 1 gal 1 wk 5.0 ab Bonide All Seasons Horticultural Spray Oil 2 gal 1 wk 5.3 a *Black spot intensity was assessed using a modified 1 to 10 Florida leaf spot rating scale. **Means followed by the same letter are not significantly different according to the least significant difference test (P<0.05). 453

36 Black Root Rot (Thielaviopsis basicola) of Ilex crenata, Ilex X Meserve and Ilex glabra in Retail Nurseries and Garden Centers A.S. Windham 1, M.T. Windham 2, L. Self 3 1 U.T. Extension, Nashville, TN U.T. Ag Research, Knoxville, TN Tennessee Department of Agriculture, Nashville, TN awindham@utk.edu Index words: Ilex, black root rot, Thielaviopsis Significance to Industry: Roots samples collected from several Ilex species at five retail nurseries and garden centers were assayed for Thielaviopsis basicola the causal agent of black root rot. At four of the garden centers infection was nearly 100% for cultivars of Ilex crenata, Ilex X Meserve and Ilex glabra. There was no consistent association of chlorotic foliage with diseased plants or healthy foliage with healthy plants. Black root rot of susceptible holly cultivars continues to be a problem in the Green Industry. Nature of Work: Ilex species are widely popular in the Green Industry due to their form, texture, evergreen foliage in many species and fruit. Unfortunately, several species are susceptible to black root rot caused by the fungus, Thielaviopsis basicola. Symptoms associated with black root rot include chlorotic foliage, branch dieback, stunting and necrotic roots (Figure 1). Black root rot has been widely reported in Ilex crenata and Ilex X Meserve cultivars in landscape plantings (1,2). The purpose of this survey was to determine the current state of black root rot in Ilex in the Green Industry in the Southeast. Root specimens were collected from Ilex species in five independent garden centers. Ilex species in the garden centers originated from large wholesale nurseries in AL, GA, SC and TN. Plants were scored for health of the foliage as either chlorotic or healthy. Three plants were chosen per cultivar; when possible, three chlorotic plants were sampled and three plants with healthy foliage were sampled. Root samples were washed free of bark media, blotted dry and placed on carrot discs placed on filter paper moistened with water in Petri plates. Plates were checked with a dissecting microscope at 20-40X (Figure 2) at 5 and 7 days for the presence of T. basicola. Results and Discussion: Black root rot was widespread in susceptible Ilex species at four of the five garden centers. Thielaviopsis was recovered from roots collected from Ilex crenata cultivars at all garden centers sampled (Table 1). Other than the widespread nature of the disease, one interesting aspect was there was no clear association of black root rot with plants with symptomatic foliage. Many plants with healthy foliage had roots that were infected with black root rot. Ilex glabra and Ilex X Meserve cultivars were also generally infected with black root rot. Ilex cornuta Carissa and dwarf Ilex vomitoria were healthy and free of Thielaviopsis. 454

37 Clearly, there is a close association of black root rot with several Ilex species. This disease has been associated with declining Ilex species in landscape beds. However, the presence of Thielaviopsis in an Ilex root system on a container plant in a garden center doesn t automatically mean that the plant will decline in a landscape bed. It is possible that cultural practices could make a difference in the future health of an infected plant. Research is needed to determine if infected plants are acceptable or unacceptable for landscape use and to determine specific cultural practices that could minimize damage from black root rot. Literature Cited: Lambe, R. C. and W. H. Ridings Black root rot of Japanese holly. Fla. Dept. Agric. & Consumer Serv. Div. of Plant Ind. Plant Path Cir. No Wick, R. L Occurrence of Thielaviopsis basicola and phytopathogenic nematodes on healthy and declining landscape-grown Ilex crenata Helleri. J. of Environ. Hort. 5(3):

38 Table 1. Black root rot incidence in Ilex species located in independent garden centers. Garden Center A Ilex species Chlorotic foliage (Y/N) % Plants with infected roots Ilex crenata Soft Touch N 100 Ilex crenata Compacta N 100 Ilex crenata Sky Pencil N 100 Ilex X Meserve Castle Spire N 100 Ilex glabra N 100 Ilex cornuta Needlepoint N 33 Garden Center B Ilex species Chlorotic foliage (Y/N) % Plants with infected roots Ilex crenata Hoogendorn N 100 Ilex crenata Soft Touch N 100 Ilex crenata Sky Pencil N 100 Ilex glabra Chamzin N 100 Ilex X Willemer N 100 Garden Center C Ilex species Chlorotic foliage (Y/N) % Plants with infected roots Ilex crenata Helleri Y 66 Ilex crenata Helleri N 100 Ilex glabra Densa N 100 Ilex crenata Soft Touch N 100 Ilex crenata Green Luster Y 100 Ilex crenata Sky Pencil N 100 Ilex crenata Hoogendorn Y 100 Ilex X Meserve Blue Princess Y 100 Garden Center D Ilex species Chlorotic foliage (Y/N) % Plants with infected roots Ilex crenata Soft Touch Y 0 Ilex crenata Soft Touch N 0 Ilex crenata Helleri N 66 Ilex crenata Compacta N 100 Ilex crenata Green Luster N 66 Ilex cornuta Carissa N 0 Ilex crenata Sky Pencil N 0 Ilex vomitoria dwarf N 0 Ilex crenata Hoogendorn N 0 Ilex glabra N 0 Ilex X Meserve Blue Princess N 0 Garden Center E Ilex species Chlorotic foliage (Y/N) % Plants with infected roots Ilex crenata Soft Touch N 66 Ilex crenata Soft Touch Y

39 Figure 1. Necrotic roots of Ilex crenata Sky Pencil infected with black root rot. Figure 2. Thielaviopsis basicola colonies growing from diseased Ilex roots on carrot discs at 7 days. 457

40 First Report of Canna Yellow Mottle Virus in Tennessee A.S. Windham 1, M.T. Windham 2, L. Self 3 1 U.T. Extension, Nashville, TN U.T. Ag Research, Knoxville, TN Tennessee Department of Agriculture, Nashville, TN awindham@utk.edu Index words: canna, virus, canna yellow mottle virus Significance to Industry: Canna yellow mottle virus (CaYMV) was first detected in Canna Bengal Tiger in Tennessee in 2008 in an independent garden center, and in Canna Tropicana Gold and Tropicana Phaison in 2009 in big box stores. Nature of Work: Cannas are herbaceous plants with colorful foliage and flowers that are widely used in the Green Industry. Many of the more recently developed cultivars, such as Bengal Tiger, Tropicana Gold and Tropicana Phaison have striking foliar variegation. In 2008, Canna Bengal Tiger with necrotic streaks in leaves was observed in a garden center in Nashville, TN. Foliage was collected and shipped to Agdia, Elkhart, IN. The leaves were assayed for CaYMV using polymerase chain reaction (PCR). The specimen was positive for CaYMV. In 2009, numerous specimens of Canna cultivars Tropicana Gold (Figures 1 and 2) and Tropicana Phaison were observed with stunted growth and necrotic streaks in the foliage. Foliar specimens of each cultivar were collected and submitted to Agdia to be assayed for CaYMV. Each cultivar was positive for the virus. Infected plants originated in large wholesale nurseries in TN and GA. Infection levels at some stores appeared to be nearly 100%. As nearly all of the plants observed in 2009 were infected, store employees were unaware of the problem, as there were no vigorous, healthy plants with normal variegation to use as a comparison with the diseased canna. Results and Discussion: CaYMV has been reported in several states in the U.S. and in several countries. In the U.S., it appears to be widespread in the nursery trade. Green industry employees at all levels of production and marketing should be aware of the symptoms of this disease so that infected plants are discarded before they are shipped to retail markets. Management of this virus disease is going to rely heavily on propagating only healthy plants which have been indexed for CaYMV, and rogueing virus infected plants. Literature Cited: Lockhart, B.E.L Occurrence of canna yellow mottle virus in North America. Acta Hort. (ISHS) 234:69-72 Momol, M. T., B. E. L. Lockhart, H. Dankers, S. Adkins Plant Health Progress. S. Yamashita et al Ann. Phytopathol. Soc. Jpn. 51:

41 Figure 1. Canna Tropicana Gold leaf with characteristic necrotic streaks indicating the presence of CaYMV. Figure 2. Canna Tropicana Gold plants infected with CaYMV (two plants on left), healthy (on right). 459

42 A Preliminary Screening of Roses for Resistance to Foliar Leaf Spots J. Mynes 1, A. Windham 2, Y. Li 1, C. Pounders 3, J. Spiers 3 and M. Windham 1 1 University of Tennessee, Knoxville, TN Thad Cochran Southern Horticultural Laboratory, Poplarville, MS Key Words: disease resistance, black spot, Cercospora leaf spot, Diplocarpon rosae, Cercospora rosicola Significance to Industry: In this preliminary study, 13 rose cultivars were found with acceptable levels of resistance to both black spot and cercospora leaf spot. However, resistance claims in catalogs of companies selling roses were not substantiated for many rose cultivars. Caution should be used in purchasing roses based on these claims. Nature of Work: The popularity of roses as an ornamental plant is tarnished because of foliar diseases that often can not be controlled without preventive fungicidal sprays applied at weekly intervals. A large percentage of gardeners avoid roses due to an aversion of conforming to a spray routine or they do not want pesticides in their gardens or stored in their garages. This aversion to pesticides contributes to the popularity of disease resistant roses, such as the extremely popular cultivar Knock Out. Rose companies have recognized the popularity of disease resistant roses and have used colorful terms to imply disease resistance. Some examples of terms used to describe resistant roses in wholesale catalogs are: resistant foliage, remarkable disease resistance, care free, trouble free, and resists the dreaded fungus. Unfortunately, retail nurseries and their customers, desiring disease resistant roses; have been disappointed when disease resistance claims could not be substantiated. In 2009, a no-spray rose trial was conducted at the West Tennessee Research and Education Center (WTREC) in Jackson, TN and the Plateau Research and Education Center (PREC) in Crossville, TN, a total of 114 cultivars were evaluated. Cultivars were included if they had been described as having resistance to disease in marketing descriptions of wholesale catalogs at least once in the last five years or whose inclusion was requested by rose breeders. The test included modern shrub roses and a number of Rosa rugosa hybrids. Hybrid tea, floribunda, grandiflora, polyantha, groundcover cultivars were also represented in the evaluations. The objectives of this study were to test the validity of marketing claims concerning disease resistance and to identify rose cultivars that exhibit superior levels of resistance to black spot and cercospora leaf spot when inoculum levels of foliar pathogens are high and the environment is conducive for disease development. Roses were transplanted in late spring/early summer in a completely random design (replications = 4) with a spacing of 4 ft in a double row with 12 ft. of grass between each double row. After transplanting, plants were watered and mulched. A drip irrigation line 460

43 was installed and plants were watered as needed during the summer. Plants were fertilized with a general fertilizer once a month until late summer when plants were allowed to harden off. Pruning was used only to prevent one plant from overgrowing an adjacent plant. No fungicides were used in the study. The cultivar Peace, known for being extremely susceptible to black spot, was planted in a grid with three plants in each replication. This cultivar served as a susceptible control and as an additional source of inoculum. Plants were evaluated every two weeks from planting until frost for susceptibility to black spot and cercospora leaf spot using the following scale: 0 = no visible symptoms, 1 = < 2% of foliage diseased, 2 = < 10% of foliage diseased, 3 = < 25% of foliage diseased, 4 = < 50% of foliage diseased, 5 = > 50% of foliage diseased, and 6 = 100% of foliage diseased. Defoliation was rated on the same scale. This scale allows for the delineation between cultivars with low disease severity levels but placed very susceptible cultivars in the same category. Data were analyzed by date and by year using the Proc GLM procedure of SAS. For both day and year variables, F-values were considered significant at the 0.05 level. When a significant F-value was detected, cultivars were separated using a LSD means separation test (p=0.05). Data analyses are included by planting year. Roses were considered to have superior resistance when disease and defoliation levels were all below 2% coverage. Roses with less than 10% coverage to all disease and defoliation were considered to be moderately resistant and may need limited spraying in a landscape setting to maintain clean foliage. Results and Discussion: Black spot and cercospora were detected in early summer in Jackson and roughly four weeks later in Crossville. By August disease epidemics had reached the logarithmic growth stage at both locations. Of the 114 cultivars trialed (Table 1), thirteen were considered resistant or no-spray and eighteen were considered moderately resistant. It is important to note that this is the first year of the trial and all cultivars will be watched closely to see if performance is similar in year two. History has taught us that with the second summer s increased disease pressure some cultivars whose performance was exemplary the first year may falter or break entirely in year two. 461

44 Table 1. List of Roses Trialed Cultivar Rose Type Resistance Groundcover Shrub Groundcover Groundcover Shrub Floribunda Floribunda Shrub Floribunda Groundcover Floribunda Shrub Shrub Shrub Apricot Candy Hybrid Tea + Baby Grand Miniature Beach Blanket Groundcover + Betty White Groundcover Blushing Knock Out Shrub ++ Bolero Floribunda Bubblicious Shrub + Caramel Kisses Miniature Carefree Celebration Shrub + Carefree Marvel Shrub Carefree Spirit Shrub Chris Everet Hybrid Tea Cinco de Mayo Floribunda + Como Park Shrub Coral Cove Shrub Coral Drift Groundcover Crimson Meidiland Shrub Daddy's Little Girl Miniature Double Knock Out Shrub Dream Come True Grandiflora Drop Dead Red Grandiflora Easy Does It Floribunda 462

45 Table 1. List of Roses Trialed, continued Cultivar Rose Type Resistance 1 Easy Going Floribunda + Elle Hybrid Tea Enchanted Evening Floribunda Eternal Flame Grandiflora Fairy Meidiland Shrub Fame! Hybrid Tea Fire Meidiland Shrub ++ Firefighter Hybrid Tea Forty Heroes Hybrid Tea Fragrant Spreader Groundcover ++ Fredric Mistral Hybrid Tea George Burns Floribunda Gingerbread Man Miniature Gizmo Miniature Golden Celebration Antique + Heartbreaker Miniature High Voltage Shrub Hope for Humanity Shrub Kashmir Shrub ++ Kimberlina Floribunda Knock Out Rose Shrub ++ Lemon Drop Miniature Livin' Easy Floribunda Lovestruck Floribunda Lynn Anderson Hybrid Tea Mardi Gras Floribunda Margret Merril Floribunda Marmalade Skies Floribunda Mellow Yellow Hybrid Tea Midwest Living Shrub ++ Moje Hammarberg Shrub ++ Morden Fireglow Shrub Mother of Pearl Grandiflora Mr. Lincoln Hybrid Tea Neon Cowboy Miniature New Zealand Hybrid Tea 463

46 Table 1. List of Roses Trialed, continued Cultivar Rose Type Resistance 1 Over the Moon Hybrid Tea Paprika Groundcover ++ Paradise Hybrid Tea Peace Hybrid Tea Peach Drift Groundcover + Peachy Cream Groundcover + Pink Double Knock Shrub ++ Out Pink Drift Groundcover + Pink Knock Out Shrub ++ Pink Meidiland Shrub + Pink Promise Hybrid Tea Pink Traviata Hybrid Tea Pope John Paul II Hybrid Tea Pumpkin Patch Floribunda Rabble Rouser Miniature Rainbow Knock Out Shrub Rainbow Sorbet Floribunda Red Drift Groundcover Roseberry Blanket Groundcover Ruby Ruby Miniature S-504 Groundcover ++ Scarlet Meidiland Shrub + Sevillana Shrub Shockwave Floribunda Snowcone Shrub Spanish Sunset Floribunda Spring Fever Groundcover Sunny Knock Out Shrub Sunrise Vigorusa Shrub Sweet Drift Groundcover + Tenny Bopper Miniature Therese Bugnet Shrub Traviata Hybrid Tea Watercolors Shrub Welcome Home Hybrid Tea What a Peach Shrub 464

47 Table 1. List of Roses Trialed, continued. Cultivar Rose Type Resistance 1 White Out Shrub Widow of the South Shrub ++ Wild Blue Yonder Grandiflora Winter Sunset Shrub + Yellow Jacket Shrub 1 ++ Resistant or having less that 2% infection + Moderately resistant or having less than 10% infection --- Susceptable 465

48 Effects of Temperature Stress on Solanum Host for the Spread of Soft-Rot Infection C. Gibson and A. Aziz Department of Agricultural Sciences, School of Agriculture and Consumer Sciences Tennessee State University, 3500 John A. Merritt Blvd., Nashville, TN Keywords: Altantic Potato, Norchip Potato, Pectobacterium (formerly Erwinia) carotorovum Significance to Industry: This study was performed to observe the effects of temperature stresses on the spread of soft-rot infection on Solanum host. A positive correlation between heat stress of host and infectivity by Pectobacterium for Solaneous plant was shown. These results are applicable to other Solanum species, including ornamental members of this genus. An understanding of how temperature affects the spread of Soft-Rot Infection will help in the development of management practices that can help slow the spread of soft-rot and cut the economic losses sustained by the industry. Nature of Work: The genus Solanum contains a large and diverse number of annuals and perennials commonly referred as horsenettles, lampshades and other relatives. These plants can grow as vines, shrubs, forbs as well as small trees while bearing attractive tubers, leaves and fruits [1]. Flowers of this genus are popular such as the Jasmine Nightshade or S. crispum [2]. However, food crops of this genus are of particular interest as being important global crops including tomato, eggplant and potato. These crops are susceptible to pectolytic bacteria causing soft-rot. Soft-rot of potatoes (Solanum tuberosum) caused by Pectobacterium (formerly Erwinia) carotorovum bacteria leads to economic loses world-wide [2]. The bacterial infection of the plant is marked by the release of large amounts of bacterial host-tissue-degrading enzymes. Eliciting plant defense response, the enzyme pectate lyase (PL) that degrades the plant cell wall pectin to oligogalacturonates, is the primary measure of bacterial virulence. This study focuses on two commercial varieties of potatoes, Atlantic (heat sensitive) and Norchip (heat tolerant) [4]. Soft-rot pathogens KD 0001 and KD 0201 are strains of P. carotorovum in which the former is naladixic acid resistant, lac z isolate and has the same virulence as the wild type and the latter being a hyper-virulent Tu5 mutant of the former (Dumenyo, unpublished). This paper investigates the effects of three levels of temperature stress on Norhcip and Atlantic tubers while these are infected with using two bacterial strains KD 0001 and KD Potato plants of the Norchip and Atlantic varieties were grown under greenhouse conditions using tuber slices as starting material. At maturity, the plant tubers were harvested and refrigerated at 10 C until use. KD 0001 and KD 0201 were mass cultured by selecting a single colony from existing plates by streaking it on media plate containing LB [5] with the antibiotics nalidixic acid and kanamycin for the former and LB and naladixic 466

49 acid for the latter. After streaking, the bacteria were allowed to grow at 28 C for forty-eight hours prior to inoculum preparation. Before infection, tubers were placed in labeled 250mL plant containers (NUNC, Kamstrupvej, Rostilde, Denmark). Three tubers were set up for each of the twelve treatments. There were four parameters for stress that were used; 28, 16, 4 and 0 hours incubation at 40 C. To prepare inoculums, the bacteria were scraped from half the surface area of the media plates with a bacterial loop and suspended in 1 ml of sterile water. For estimating the number of cells, 50μL aliquots of each suspension were used to determine the optical density (O.D.). The suspensions were diluted to an O.D. of 1 before injecting 10µL into the tubers using a micropipette. For the controls, water was used instead of bacterial suspension. After infection, the tubers were placed in a 28 C incubator and were observed over a five day period before macerated (rotted) tissue was extracted and weighed. Results and Discussion Results from this study showed that there is a significant difference in the level of soft-rot infections when tubers were exposed to either non-stressed or temperature stress conditions. The maximum infection level was observed when tubers sustained 16 hours of temperature stress and thus provided the most macerated tissue (Table 1). KD 0001 and KD 0201 showed no discernable differences in virulence in this treatment (Table 1). The Atlantic variety showed more susceptibility to infection than the Norchip variety. These results suggest the relative heat sensitivity of Atlantic [3] contributed the level of soft rot observed (Table 1); temperature stress and the spread of soft-rot infection on host are proportionally correlated to each other. Acknowledgments The authors wish to thank USDA for funding this research, Tennessee State University School of Agricultural and Consumer Sciences for the laboratory facilities used for this study. Technical help by Caleb Kersey, Carl Darris, V. Sahiti Komereddy, Paul Agyemang and Zong Liu, C. Korsi Dumenyo is also acknowledged. Literature Cited 1. Christman, Steve #680 Solanum tuberosum. Floridata. (Nov. 13, 2009) 2. The Royal British Horticultural Society Award of Garden Merit Plants 2008 with descriptions drawn up by the Plants Trials Sub-Committee Oct (Nov. 11, 2009) 3. Christina B. Wegener Induction of defence responses against Erwinia soft rot by an endogenous pectate lyase in potatoes, Physiological and Molecular Plant Pathology, 60 (2): Yeh-Jin Ahn, Kristine Claussen, J. Lynn Zimmerman Genotypic differences in the heat-shock response and thermotolerance in four potato cultivars, Plant Science, 166 (4): Maria P MacWilliams Luria Broth (LB) and Luria Agar (LA) Media and Their Uses Protocol, American Society for Microbiology, (Nov. 13,

50 Table 1. The weight of potato rotted tissue (g) after the tubers of two varieties (Atlantic and Norchip) were exposed to four stress treatments and two pathogens. Varieties Atlantic Norchip Heat Stress 0hrs 4hrs 16hrs 28hrs 0hrs 4hrs 16hrs 28hrs KD ± ± ± ± ± ± ± ± KD ± ± ± ± ± ± ± ±

51 Figure1: Effects of 40 C stress at different durations and soft-rot infection. (Pectobacterium carotorovum KD 0001 or KD 0201) on Atlantic (heat sensitive) and Norchip (heat tolerant) potatoes. A positive correlation between heat stress of host and infectivity by Pectobacterium is applicable. A to D: Atlantic potatoes infected with KD 0001 at 0, 4, 16, and 28hrs of heat stress. E to H: Atlantic potatoes infected with KD 0201 at 0, 4, 16, and 28hrs of heat stress. I to L: Norchip potato infected with KD 0001 at 0, 4, 16, and 28hrs of heat stress. M to P: Norchip potato infected with KD 0201at 0, 4, 16, and 28hrs of heat stress. 469